Intestinal microbiota and enteric alterations in Poecilia reticulata exposed to environmentally relevant azithromycin concentrations

The intestinal microbiota is essential for host homeostasis, influencing immune function, metabolism, nutrient absorption, and inflammatory regulation. In aquatic ecosystems, chronic exposure to antibiotics through continuous discharge can disrupt the microbial communities of non-target organisms, compromising intestinal health. This study investigated how environmentally relevant concentrations of azithromycin (AZM), a widely prescribed macrolide antibiotic, alter the intestinal microbiota and homeostasis of the guppy ( Poecilia reticulata ). Fish were exposed to 2, 4, and 16 µg/L of AZM for 15 days and subsequently euthanized for intestinal analysis. Exposure to AZM promoted exploratory changes in the intestinal microbial community composition and diversity profiles across exposure groups. These alterations were accompanied by histological and cellular changes, including increased goblet cell and intraepithelial lymphocyte densities and variations in intestinal wall thickness. The density of vasoactive intestinal peptide (VIP)-positive neurons decreased, while nitrergic neurons (NADPH-diaphorase positive) increased, suggesting changes in neuronal density. Furthermore, azithromycin exposure altered lipid peroxidation levels in the gastrointestinal tract, with increased oxidative damage observed at the highest concentration tested, whereas DNA integrity remained unaffected. Collectively, these results suggest that environmentally relevant concentrations of azithromycin may alter microbial composition alongside intestinal and enteric neuronal populations in P. reticulata . Such disturbances could potentially affect intestinal physiological regulation. These findings highlight the ecological risk posed by chronic antibiotic contamination in aquatic environments.